Origen Therapeutics is committed to developing pharmaceutical products in transgenic chickens. In particular, Ongen's mission is to develop high affinity human antibodies. by creating chickens in which the endogenous heavy and light chain immunoglobulin (Ig) genes are 'knocked-out' and reconstituted with the human heavy and light chain Ig genes. In evolutionary terms, birds have diverged from mammals and, therefore, the chicken produces high affinity antibodies against conserved mammalian antigens. In addition, chickens produce a wide repertoire of high titer antibodies that possess high affinity and avidity. By knocking out the endogenous lg loci and inserting the human loci in germline configuration, human polyclonal antibodies can be isolated from the chicken's egg. By conventional techniques, human monoclonal antibodies can be produced from B-cells derived from chickens carrying inactivated endogenous Ig loci and human Ig loci in germline configuration. These chickens will provide unique access to human polyclonal antibodies as therapeutic agents. Human poylclonal antibodies are desirable because they bind to several epitopes on the target antigen, making them highly effective against complex antigens such as bacteria, fungi, viruses and toxins. Currently, human polyclonal antibodies are used as anti-infectious agents against a variety of diseases but commercially available immunoglobulin preparations from human donors vary in their titer and effectiveness for pathogens, severely limiting their utilization. In spite of their attractiveness as therapeutics, high titer human antibodies from immunized sources are not available.Origen's technology for producing chickens carrying the human Ig repertoire is based on the use of chicken embryonic stem cells. This application proposes to develop methods to specifically delete the chicken IgH and lgL chain genes. The first aim is to develop robust methods for transfecting and selecting cES cells and to show germline transmission of a transgene. The second aim is to target the J-segments of the heavy and light chain loci to prevent translation of the endogenous genes. Correctly targeted ES cells will be injected into recipient embryos to produce germline chimeras. Heterozygous lgHKO and lgLKO birds will be produced and will be bred in phase Ill to birds carrying the human IgH and lgL genes. At the completion of phase Ill, chickens producing human Ig in response to antigen will yield a novel class of therapeutic proteins providing highly effective passive immunity to patients suffering from acute diseases for which therapy is not currently available and chronic diseases that require specific, high titer polvclonal antibodies.